Chronic hypersensitive pain states can become established following sustained,
repeated or earlier noxious stimuli and are notably difficult to treat, especially in
cases where nerve injury contributes to the trauma. A key underlying reason is that a
variety of plastic changes occur in the central nervous system (CNS) at spinal and
potentially also supraspinal levels to upregulate functional activity in pain processing
pathways. A major component of these changes is the enhanced function of
excitatory amino acid receptors and related signalling pathways.
Here we utilised rodent models of neuropathic and inflammatory pain to investigate
whether evidence could be found for lasting hypersensitivity following neonatal (or
adult) noxious stimuli, in terms of programming hyper-responsiveness to subsequent
noxious stimuli, and whether we could identify underlying biochemical mechanisms.
We found that neonatal (postnatal day 8, P8) nerve injury induced either long lasting
mechanical allodynia or shorter lasting allodynia that nonetheless was associated
with hyper-responsiveness to a subsequent noxious formalin stimulus at P42 despite
recovery of normal mechanical thresholds. By developing a new micro-scale method
for preparation of postsynaptic densities (PSD) from appropriate spinal cord
quadrants we were able to show increased formalin-induced trafficking of GluA1-
containing AMPA receptors into the PSD of animals that had received (and
apparently recovered from) nerve injury at P8. This was associated with increased
activation of ERK MAP kinase (a known mediator of GluA1 translocation) and
increased expression of the ERK pathway regulator, Sos-1. Synaptic insertion of
GluA1, as well as its interaction with a key partner protein 4.1N, was also seen in
adults during a nerve injury-induced hypersensitive pain state.
Further experiments were carried out to develop and optimise a new technological
platform enabling fluorometric assessment of Ca2+ and membrane potential
responses of acutely isolated CNS tissue; 30-100 μm tissue segments,
synaptoneurosomes (synaptic entities comprising sealed and apposed pre- and postsynaptic
elements) and 150 × 150 μm microslices. After extensive trials, specialised
conditions were found that produced viable preparations, which could consistently
deliver dynamic functional responses. Responsiveness of these new preparations to
metabotropic and ionotropic receptor stimuli as well as nociceptive afferent stimulant
agents was characterised in frontal cortex and spinal cord.
These studies have provided new opportunities for assessment of plasticity in pain
processing (and other) pathways in the CNS at the interface of in vivo and in vitro
techniques. They allow for the first time, valuable approaches such as microscale
measurement of synaptic insertion of GluA1 AMPA receptor subunits and ex vivo
assessment of dynamic receptor-mediated Ca2+ and membrane potential responses
